In vitro toxicity testing of phthalocyanines on different cell lines using a continuous laser source

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Odhav, Bharti

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Karsten, Aletta

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Maduray, Kaminee

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2010-11-17T13:31:38Z

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2012-09-01T22:20:06Z

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2010

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331449

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http://hdl.handle.net/10321/550

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Submitted in fulfilment of the requirements of the Degree of Master of Technology: Biotechnology, Durban University of Technology, 2010.

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Photodynamic therapy is a promising treatment for cancer. It involves the
combination of a photosensitizer and light of an appropriate wavelength (laser source)
to cause the destruction of cancer cells. Phthalocynanines are second–generation
photosensitizers with enhanced photophysical and photochemical properties.
In this in vitro study the effect of aluminium (AlTSPc) or zinc (ZnTSPc)
tetrasulfophthalocyanines in its inactive and active state (laser induced) on melanoma
(skin cancer cells), fibroblast (healthy normal skin cells) and keratinocyte (healthy
normal skin cells) cells was evaluated. For each of the cell lines approximately
3 x 104 cells/ml were seeded onto 24-well cell culture plates and allowed to attach
overnight, after which cells were treated with different concentrations of AlTSPc or
ZnTSPc. The photosensitizers were synthesized at Rhodes University. After 2 hrs,
cells were irradiated with a diode laser at a wavelength of 672 nm and a beam
diameter of 1 cm. The laser power varied between 20-30 mW and the irradiation time
was calculated to deliver a light dose of 4.5 J/cm2. Post-irradiated cells were
incubated for 24 hrs before cell viability was measured using the CellTiter-BlueTM
Viability Assay.
Also, the efficacy of the light dose and laser source used for the killing of
approximately 50% of the melanoma cancer cells were investigated. AlTSPc and
ZnTSPc decreased cell viability of melanoma cancer cells to approximately 50% with
photosensitizer concentrations of 40 μg/ml and 50 μg/ml respectively. These
photosensitizer concentrations caused a slight decrease in the percentage cell viability
of fibroblast and keratinocyte cells. Results for the dark toxicity assay showed that
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both photosensitizers in the presence of high concentrations (60 μg/ml – 100 μg/ml)
showed cytotoxicity effects on melanoma cancer cells in their inactive state. This was
not observed in fibroblast and keratinocyte cells treated under the same experimental
conditions. The optimal AlTSPc and ZnTSPc concentrations in combination with the
light dose of 4.5 J/cm2 was the most efficient in killing the melanoma cancer cells
with reduced killing effects on healthy normal fibroblast and keratinocyte cells when
compared to other light doses (2.5 J/cm2, 7.5 J/cm2 and 10 J/cm2).
The irradiation of cells photosensitized with the optimal photosensitizer
concentrations with a femtosecond laser using similar laser parameters to continuous
wave laser experiments resulted in a reduction in the cell viability of healthy normal
fibroblast and keratinocyte cells compared to melanoma cancer cells. The presence of
DNA degradation on agarose gel, morphological changes like blebbing and
ultrastructural changes like nucleus condensation indicated that photodynamic therapy
treated melanoma cancer cells with the optimal concentrations of AlTSPc and
ZnTSPc induced cell death via apoptosis. This concludes that low concentrations of
AlTSPc and ZnTSPc activated with an appropriate laser source can be used to induce
cell death in melanoma cancer cells. Both AlTSPc and ZnTSPc exhibit the potential to
be used as a photosensitizer in photodynamic therapy for the treatment of melanoma
cancer with the occurrence of minimal damage to surrounding healthy tissue.

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136 p

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en

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Biotechnology

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Phthalocyanines

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Toxicity testing--In vitro

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Cell lines

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Lasers

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dc.title

In vitro toxicity testing of phthalocyanines on different cell lines using a continuous laser source